Interpretive Summary: Contaminated irrigation water is one of the most common vehicles by which E. coli O157:H7 may be introduced into the environment. One of the most common vehicles by which E. coli O157:H7 may be introduced into crops is by flood irrigation with water contaminated with cattle feces or by contaminated surface water. In this experiment, we used bacteria tagged with green fluorescent protein in a growth chamber study for easy identification of bacterial survival in soil, roots and leaf surfaces. Soil samples and plant materials were collected weekly. After plating and PCR analysis of these samples, E. coli O157:H7 concentrations were greater on the rhizosphere than the non-rhizosphere soils and leaf surfaces. E. coli O157:H7 survived longer than 45 days in both rhizosphere and soil samples. Our results showed that PCR analysis was faster and more reliable in enumerating the pathogen than plate counts.

Technical Abstract:
Real-time PCR method was developed to detect and quantify Escherichia coli O157:H7/pGFP (E. coli). Probe was designed to hybridize with the eae gene of E. coli O157:H7. The probe was incorporated into real-time PCR containing DNA extracted from the phyllosphere, rhizosphere, and non-rhizosphere soils irrigated with water artificially contaminated with E.coli O157:H7. The detection limit for E. coli O157:H7 quantification by real-time PCR was 1.4 x 103 in rhizoshere and phyllosphere samples. E. coli O157:H7 concentrations were higher on the rhizosphere than in the non-rhizosphere soils and leaf surfaces, and persisted longer in clay soil. The concentrations of E. coli O157:H7 obtained by real-time PCR were comparable to the concentrations obtained by traditional culture methods during the first three days of inoculation, and thereafter, the concentrations by real-time PCR were higher. The persistence of E. coli O157:H7 in phyllosphere, rhizosphere, and non-rhizosphere soils over 45 days may play a significant part in the re-contamination cycle in the environment. Therefore, the rapidity and the feasibility of the assay may be a useful tool for quantification and monitoring of E. coli O157:H7 in irrigation water.